Pref-1 (Preadipocyte factor-1) is expressed in adipose precursor cells, downregulated during differentiation and is absent in mature adipocytes. Thus, Pref-1 is widely used as a preadipocyte marker. We have shown Pref-1 Inhibition of adipogenesis by gain- and loss- of function studies in vitro as well as in vivo. Pref-1 is synthesized as a plasma membrane glycoprotein with 6 EGF-repeats in its extracellular domain; the Pref-1 ectodomain is cleaved by TACE at the juxtamembrane region to generate the biologically active soluble Pref-1. Pref-1 inhibits adipocyte differentiation by activating MEK/ERK and we recently detected an interaction of Pref-1 with fibronectin. The goal of this research is to further elucidate the molecular mechanisms underlying the Pref-1 function in adipose tissue development. First, we will examine the Pref-1 interaction with fibronectin and the specific integrin receptor that mediates Pref-1 inhibition of adipocyte differentiation. We will also study the fibronectin/integrin signaling pathway that is dependent on the Pref-1/fibronectin interaction in inhibiting adipocyte differentiation. Downstream signaling molecules including FAK and Rac will be examined. Next, to mark and characterize Pref-1 expressing adipose precursor cells, we will generate Pref-1- reporter mice that express GFP and RFP in an inducible manner. Using these reporter mice, we will monitor and examine Pref-1 expressing adipose precursor cells. We will also define the signature of these cells by genome-wide microarray analysis. Furthermore, comparison of the expression patterns as affected by Pref-1 null and Pref- 1 overexpression will help us to understand the Pref-1 function in adipose precursor cells.